Complement lysis of erythrocytes (E) is mediated by C5-C9, inserting into lipid bilayers of cell membranes. We reported another mode of C5 activation which involves acidification to pH 6.4 of a mixture of C5 and C6. In the presence of C7, C8 and C9, the mixture, designated C(56)a generates a lytic activity for chicken E. We earlier performed a comparative study of the biological behavior of C(56)a and C5b,6 and now have extended these studies on the functional and physico-chemical properties of C(56)a. The acid generation of C(56)a is dose dependent on C5 and C6 and is as efficient as classical or alternative pathway activation. While C(56)a decays in the presence of C7, the acid activated complex, is also labile at 37 C in the absence of C7. Upon ultracentrifugation, acid activated C5 and C6 sediment consistent with the formation of a bimolecular 11S complex that coincides with the appearance of lytic activity. Analysis of the chain structure of C5 obtained from C(56)a prepared from purified components or from Egp ghosts demonstrates a C6 dependent, C5 chain cleavage. Studies on the interaction of acid-activated terminal C5b,6a-9 with paroxysmal nocturnal hemoglobinuria (PNH)-E and NHE indicates that C5b, 6a appears to be a principal lytic factor in acidified human serum.